Terephthalic acid and isophthalic acid are two monomers having utility in the production of polyesters which are commercially required in large quantities for fibers, films, paints, adhesives and beverage containers. Isophthalic acid is also used in condensation reactions with diamines to form polyamides. Polyesters and polyamides are two extremely important classes of commercial polymers.
A variety of chemical routes to terephthalic acid and isophthalic acid are known. The most notable commercial process to prepare terephthalic acid involves the liquid-phase oxidation of p-xylene. The Amoco process involves oxidizing p-xylene with a molecular oxygen-containing gas in the liquid phase in a lower aliphatic monocarboxylic acid solvent in the presence of a heavy metal catalyst and a bromine compound to from terephthalic acid directly (U.S. Pat. No. 2,833,816). More specifically, the reaction is catalyzed by Co and Mn in 95% acetic acid with a mixture of NH.sub.4 Br and tetrabromoethane as cocatalysts. The oxidation is carried out under severe conditions of high temperatures (109-205.degree. C.) and pressures (15-30 bar). Hence, the rate of reaction is high and the yield of terephthalic acid based on p-xylene is as high as 95% or more. However, the reaction apparatus becomes heavily corroded owing mainly to the use of the bromine compound and the monocarboxylic acid solvent. Thus, ordinary stainless steel can not be used to build the reaction apparatus, and expensive materials such as Hastelloy or titanium are required. In addition, because the acid solvent is used in large quantity and the oxidation conditions are severe, combustion of the solvent itself can not be avoided, and its loss is not negligible. The Amoco process has also been shown to oxidize m-xylene to isophthalic acid. Although it is possible to oxidize xylenes by these methods, they are expensive and generate waste streams containing environmental pollutants.
Biological oxidation of methyl groups on aromatic rings, such as toluene and isomers of xylene, is well known (Dagley et al., Adv. Microbial Physiol. 6:1-46 (1971)). For example, bacteria that have the xyl genes for the Tol pathway sequentially oxidize the methyl group on toluene to afford benzyl alcohol, benzaldehyde and ultimately benzoic acid. The xyl genes located on the well characterized Tol plasmid pWWO have been sequenced (Assinder et al., Adv. Microbial Physiol. 31:1-69 (1990); Burlage et al., Appl. Environ. Microbiol. 55:1323-1328 (1989)). The xyl genes are organized into two operons. The upper pathway operon encodes the enzymes required for oxidation of toluene to benzoic acid. The lower pathway operon encodes enzymes that convert benzoic acid into intermediates of the tricarboxylic acid (TCA) cycle.
In addition to toluene, m-xylene and p-xylene are substrates of the Tol pathway (JP 9023891). The upper pathway enzymes catalyze oxidation of one methyl group on m-xylene and p-xylene to produce the corresponding methylbenzyl alcohol, methylbenzaldehyde and methylbenzoic acid. Although many bacteria utilize m-xylene and/or p-xylene as sources of carbon and energy for growth, essentially all of the known examples oxidize m-xylene or p-xylene to methylbenzoic acid (i.e., m-toluic acid or p-toluic acid) and then convert the methylbenzoic acid to methylcatechol. Bacteria with the Tol pathway have not been shown to produce isophthalic acid or terephthalic acid as intermediates when p- and m-xylene are used as substrates. However, certain bacteria are known to oxidize the methyl group of methylbenzoic acid when this compound is degraded to provide carbon and energy for growth. For example, Comamonas testosteroni strain T-2 oxidizes p-methylbenzoic acid (p-toluic acid) to terephthalic acid (Junker et al., J. Bacteriol. 179:919-927 (1997); Junker et al., Microbiology 142:2419-2427 (1996)). It is important to note that although this strain degrades methylated aromatics such as p-toluenesulfonic acid, it displays no activity against toluene or p-xylene.
In general, biological processes for production of chemicals are desirable for several reasons. One advantage is that the enzymes that catalyze biological reactions have substrate specificity. Accordingly, it is sometimes possible to use a starting material that contains a complex mixture of compounds to produce a specific chiral or structural isomer via a biological process. Another advantage is that biological processes are commonly perceived as being less harmful to the environment than chemical manufacturing processes. These advantages, among others, make it desirable to use p-xylene or m-xylene as the starting materials for manufacture of terephthalic acid or isophthalic acid, respectively, by means of a bioprocess.
SU 419509 claims a method for the cooxidative production of terephthalic acid by the microbiological transformation of p-xylene using an active culture from the genus Nocardia, which carries out the direct oxidation of p-xylene to terephthalic acid. The method is described as a cooxidative process that involves providing the bacteria with hexadecane and p-xylene. Since this is a cooxidative process, the hexadecane is required to induce synthesis of the appropriate oxidative enzymes.
Finally, JP 9023891 claims a method for the production of aromatic carboxylic acids by oxidation of various aromatic compounds by Mycobacterium sp. strain NS12523 and similar bacteria belonging to the genus Mycobacterium. Terephthalic acid and isophthalic acid are two of the aromatic carboxylic acids claimed to be formed by the described process. However, there was no demonstration that the claimed Mycobacterium strains could actually produce terephthalic acid or isophthalic acid. When p-xylene was used as a substrate, only p-toluic acid was isolated as the product. If p-toluic acid could be used as a substrate, as was claimed, it would be reasonable to expect terephthalic acid to be isolated with or as the final product.
A need exists for environmentally friendly, safe and economical methods to produce compounds of commercial interest. A method that has broad applicability for the production of terephthalic acid and isophthalic acid would have great commercial value. To the best of applicants' knowledge, there is no account of any bacteria that can oxidize both methyl groups of p-xylene to form terephthalic acid in the absence of a cooxidized compound. Furthermore, no such method involving the biological oxidation of both methyl groups of p- and m-xylene by a single organism was previously known.